(1) Field of the Invention
The present invention generally relates to regulation of kinase activity. More specifically, the invention is directed to inhibitors of AbI kinases.
(2) Description of the Related Art
Nonreceptor tyrosine kinases are major regulators of cell signaling downstream of extracellular stimuli. The ubiquitous nonreceptor tyrosine kinase c-AbI tyrosine kinase has been implicated in cell proliferation, apoptosis downstream of growth factors, integrin, and cytokine signaling (Woodring et al., 2003). The critical role of c-AbI kinase in cell proliferation is illustrated by the manifestation of chronic myelogenous leukemia (CML) due to expression of BCR-AbI, a kinase-activated mutant form of c-AbI (Goldman and Melo, 2003). However, it is not clear how BCR-AbI or c-AbI kinase activity is regulated in cells.
Auto-inhibition has emerged as the major mechanism of regulation of nonreceptor tyrosine kinases such as c-Src and c-AbI (Courtneidge, 2003; Hantschel et al., 2003; Nagar et al., 2003; Thomas and Brugge, 1997; Wang, 2004). These kinases share high structural homology conferred by the presence of highly conserved structural domains: SH3, SH2, and the catalytic domain. Crystal structures of c-Src (Williams et al., 1997; Xu et al., 1999) and c-AbI (Hantschel et al., 2003; Nagar et al., 2003) reveal that their SH3 and SH2 domains bind to the catalytic domain and induce an autoinhibitory conformation, providing the basic mechanism of regulation of these kinases. Additional regulation of c-Src and c-AbI is, however, provided by nonhomologous sequences of these kinases imposing additional inhibitory constraints. In c-Src this inhibition is achieved by intramolecular interaction of the SH2 domain with the phosphorylated tyrosine 527 located in C-terminus on the same molecule (Thomas and Brugge, 1997). In c-AbI there is no internal phosphotyrosine-SH2 domain interaction, precluding this inhibitory mechanism. In further contrast to c-Src, additional inhibitory constraints are imposed onto c-AbI by the myristoylated cap binding directly to the C-terminal lobe of the kinase domain thus further locking an SH3-SH2 “clamp” onto the catalytic domain. This “molecular lock”, however, does not exist in the nonmyristoylated form of c-AbI—1a, which contains only a partial cap region. A more comprehensive role of the cap region could not be discerned from the crystal structure of c-AbI, but the cap region is believed to play a role in regulating accessibility and binding of phosphotyrosine-containing peptides (Hantschel et al., 2003).
Various proteins that bind to c-AbI kinase have been proposed to be c-AbI co-inhibitors. The proteins F-actin, Pag and Rb have been hypothesized to act as passive co-inhibitors by stabilizing the auto-inhibited conformation of c-AbI (Wang, 2004). Another set of candidate c-AbI inhibitors, Abi1 and Abi2, were isolated in the search to identify c-AbI-binding partners (Dai and Pendergast, 1995; Shi et al., 1995) as well as in efforts to identify binding partners of the SH3 domain from other proteins, hence their names E3b1 (eps8 SH3 binding protein 1—Biesova et al., 1997) and Hssh3bp1 (human spectrin Src homology 3 domain binding protein 1—Ziemnicka-Kotula et al., 1998). The inhibitory role of Abi proteins in c-AbI kinase signaling has been proposed primarily based on the Abi1 and Abi2 inhibitory role in regulation of cell growth (Dai and Pendergast, 1995; Macoska et al., 2001; Shi et al., 1995), but the molecular mechanism of their action is not clear. Abi1 and Abi2 interact with c-AbI kinase C-terminal PXXP sequences in the proline rich linker, PRL (Dai and Pendergast, 1995; Shi et al., 1995) and with c-AbI SH3 domain (Ziemnicka-Kotula et al., 1998). Thus, one such mechanism might involve reinforcement of the c-AbI kinase autoinhibited conformation through cooperative binding of the Abi's PXXP sequences and SH3 domain to the c-AbI SH3 domain and PRL region. However, no SH2-based mechanism of kinase regulation has been demonstrated for c-AbI kinase and Abi proteins.
Based on the above discussion, characterization of the role of Abi proteins in c-AbI inhibition is needed. The present invention addresses this need.